Metatranscriptomic evidence for classical and RuBisCO-mediated CO2 reduction to methane facilitated by direct interspecies electron transfer in a methanogenic system

In a staged anaerobic fluidized-bed ceramic membrane bioreactor, metagenomic and metatranscriptomic analyses were performed to decipher the microbial interactions on the granular activated carbon. Metagenome bins, representing the predominating microbes in the bioreactor: syntrophic propionate-oxidizing bacteria (SPOB), acetoclastic Methanothrix concilii, and exoelectrogenic Geobacter lovleyi, were successfully recovered for the reconstruction and analysis of metabolic pathways involved in the transformation of fatty acids to methane. In particular, SPOB degraded propionate into acetate, which was further converted into methane and CO2 by M. concilii via the acetoclastic methanogenesis. Concurrently, G. lovleyi oxidized acetate into CO2, releasing electrons into the extracellular environment. By accepting these electrons through direct interspecies electron transfer (DIET), M. concilii was capable of performing CO2 reduction for further methane formation. Most notably, an alternative RuBisCO-mediated CO2 reduction (the reductive hexulosephosphate (RHP) pathway) is transcriptionally-active in M. concilii. This RHP pathway enables M. concilii dominance and energy gain by carbon fixation and methanogenesis, respectively via a methyl-H4MPT intermediate, constituting the third methanogenesis route. The complete acetate reduction (2 mole methane formation/1 mole acetate consumption), coupling of acetoclastic methanogenesis and two CO2 reduction pathways, are thermodynamically favorable even under very low substrate condition (down to to 10(-5) M level). Such tight interactions via both mediated and direct interspecies electron transfer (MIET and DIET), induced by the conductive GAC promote the overall efficiency of bioenergy processes.